Angiogenesis is essential for breast cancers to grow beyond minimal size. The prognosis of invasive breast cancer has been correlated with tumor vascularity, and angiogenesis may mark early progression toward malignancy in that some pre-invasive breast cancers have already induced new blood vessels. Despite its importance, the pathogenesis of tumor angiogenesis is poorly understood. Neoplastic cells are thought to initiate angiogenesis by secreting cytokines that stimulate endothelial cells (ECs) to divide and migrate and that provide an altered extracellular matrix which favors BC migration. Vascular permeability factor (VPF), also known as vascular endothelial growth factor (VEGF) is a tumor-secreted cytokine likely to have an important role in tumor angiogenesis. VPF/VEGF is a selective BC mitogen that renders microvessels hyperpermeable to circulating macromolecules. Recent studies have shown that VPF/VEGF is overexpressed in many primary human breast cancers, including both invasive and in situ ductal carcinomas. Also, two high affinity VPF/VEGF receptor tyrosine kinases, flt-1 and kdr, are overexpressed in human breast cancer BCs. The experiments proposed here will test four specific hypotheses: l. that VPF/VEGF has a central role in breast cancer angiogenesis. 2. that flt- 1 and/or kdr mediate VPF/VEGF's angiogenic effect. 3. that upregulation of VPF/VEGF receptors enhances EC responsiveness to VPF/VEGF and 4. that cytokines yet to be identified modulate tumor angiogenesis by increasing the expression of one or both VPF/VEGF receptors. More specifically, Aim 1 will evaluate the expression of VPF/VEGF, its receptors, and other cytokines and quantitate angiogenesis in human breast lesions resected at various stages of malignant progression. Aim 2 will extend these studies to breast/mammary carcinomas growing in immunodeficient mice; tumor cells will be transfected so as to express sense, anti sense or mutant VPF/VEGF. VPF/VEGF expression will then be directly related to angiogenesis, supplementing the human studies of Aim 1. Aim 3 will investigate the importance of flt-1 and kdr in mediating VPF/VEGFs several effects, including angiogenesis, using specific antibodies that selectively block one or the other receptor. Finally, Aim 4 will investigate the role that cytokines, and possibly estrogen/progesterone, play in causing overexpression of both VPF/VEGF receptors as is characteristic of breast cancer; they will also determine whether receptor overexpression alters EC responsiveness to VPF/VEGF. Collectively, these experiments will test several important hypotheses, generate fundamentally important new insights into the mechanisms by which breast (and presumably other) cancers induce and regulate angiogenesis and suggest new approaches to therapy.